Urticaria is a common skin disorder involving mast cell activation and degranulation. Urticaria is classified according to its chronicity into acute and chronic forms. It may occur spontaneously or upon exposure to a physical factor. In the latter case, the urticaria is classified as a physical urticaria. Physical urticaria may be induced by mechanical and applied pressure, exercise, or exposure to cold, heat, sun, water, or vibration. The pathologic basis of physical urticarias in general remains unclear and a genetic basis for these disorders is only partly understood. The purpose of this project is to investigate the mast cell dependent pathogenic mechanisms of physical urticaria, both to better understand how to manage urticarial inflammation and to explore the consequences of mast cell degranulation in human tissues. In these studies, patients undergo standard challenge testing to verify their urticaria. Blood samples are obtained for the investigation of molecular and genetic pathways involved in the disease process. Following the clinical induction of urticarial manifestations, additional blood samples are collected to determine soluble mediators involved in pathogenesis. Photographic imaging studies may be performed during challenge testing. Skin biopsies may be obtained prior to and following challenge testing and are analyzed for biochemical and histological markers. Since the inception of the physical urticaria protocol in 2009, we have enrolled over 160 patients including 25 healthy subjects. All patients safely underwent challenge testing based on their history. Blood samples and skin biopsies have been collected and stored for biochemical, molecular and, when applicable, genetic analysis. Mast cell degranulation was verified by skin biopsy. The majority of the patients were challenge positive to either cold-induced, cholinergic, dermatographism, solar or vibratory urticaria. In fiscal year (FY)2016, through whole exome sequencing we identified a missense mutation (p.C492Y) in the adhesion G-protein coupled receptor ADGRE2 as the genetic bases of vibratory urticaria. This finding marked the first identification of a genetic basis for a mast cell-mediated hereditary urticaria induced by a mechanical stimulus and suggests that ADGRE2 subunit interactions may also have implications for other diseases involving mast cells (NEJM Feb 2016). To further elucidate mechanisms by which interactions with ADGRE2 leads to vibratory urticaria, in FY 2019 we have shown that human mast cells expressing p.C492Y-ADGRE2 exhibit increased degranulation and cytosolic calcium flux upon vibratory mechanical stimulation and these events are required for downstream signaling and cytokine production. Our studies provide insights into the perplexing chain of molecular events triggered by mechanical forces via ADGRE2 and identify possible therapeutic targets for patients with vibratory urticaria. Other patients and families affected with vibratory urticaria are being evaluated and screened for novel mutations in ADGRE2. Subsequent studies are underway in the Mast Cell Biology Section (MCBS) to further elucidated the mechanism of ADGRE2 receptor activation and signaling in patients with inherited vibratory urticaria. In FY 2019, we published a study of exercise induced mast cell activation in patients with indolent systemic mastocytosis (ISM) compared to those with cholinergic urticaria and normal volunteers. We provided evidence that patients with ISM exhibit an intercorrelated rise in both serum histamine and tryptase follow exercise, whereas in those patients with cholinergic urticaria only an elevation in histamine was detected. Our current study corroborates with an accumulating body of evidence that suggests that physical exercise can trigger mast cell degranulation (J ALLER CL IMM-PRACT 2019). This study also suggests that clinicians managing patients with physically induced mast cell mediated manifestations should provide instructions regarding the effects of exercise and guidance in the use of mast cell mediator receptor antagonists that may offer relief of symptoms, as well as the self-administration of epinephrine for the treatment of anaphylaxis should it occur. Increased reactivity to dermal mast cells in patients with physical urticaria was contrasted with patients with GATA2-deficient subjects which exhibit evidence of impaired IgE mast cell activation and reduced IgE mediated clinical allergic disease. In this rare disease, despite conserved numbers of skin mast cells, there was decreased expression of KIT, the primary receptor for mast cell proliferation (J ALLER CL IMM 2019).